Hey everyone! My name is Jennifer Ye and I'm from Toronto, Canada. I am going into grade 11 (also called Junior Year, which I've learned from the short time I've been here) in the fall, and absolutely love learning new things. My hobbies include fencing (epee), watching House M.D., playing the violin, playing video games (Pokemon and Fire Emblem are some of my favourites!), going out with my friends, and volunteering. At school, I am the Concertmaster of my school Orchestra, Piano Accompanist for my school choir, and the Co-Chair of the Sick Kids Foundation Club and Anime Club. In my community, I volunteer and perform with my local orchestra.
When I was in Elementary School, I remembered hating Biology and dreading to go to science class. However, after a course in Grade 9 Science with an awe-inspiring teacher who majored in Biology, I learned to love everything from plants to genetics, and was hooked! I'm hoping to major in Biology when I graduate, but I'm still unsure whether I want to attend University in Canada or in the U.S., as both are very worthy contenders!
Iron-Oxidizing Bacteria to Detect Lyme Disease
Introduction/ Purpose A bacterial virus that affects at least 300,000 Americans a year, Lyme Disease is surprisingly still a large mystery to the public despite the rapidly increasing infection rates. It begins with a bite from a tick that latches onto the human skin. As ticks are small in size, they are often left unnoticed and undisturbed as they are feeding on humans. If the tick is left to feed for over 48 hours, the tick is more likely to transfer Borrelia burgdorferi (the type of bacteria that causes Lyme disease) to its victim; the chances of contracting the disease increases as the tick’s feeding time is prolonged.
An adult female tick. From: https://extension.umaine.edu/ipm/tickid/tick-photos/ The Borrelia burgdorferi bacteria itself is a spirochete, meaning that it has a spiral shaped body and flagella enclosed between the outer and inner membranes. Many other threatening diseases are spirochetes as well, implicating the consequences of contracting Borrelia burgdorferi. Compared to the other members of the spirochete family, the Borrelia burgdorferi is rather small, and its flagella is unique as it is hidden beneath the outer membrane instead of it being external. In addition, it does need iron to go through the electron transport chain to produce energy, and also has a slow reproductive rate, tricking the body into taking a longer time to develop antibodies to defend itself.
Early symptoms of Lyme disease include a bull’s-eye rash, a definitive indicator of Lyme disease, however, many do not have such obvious symptoms and develop more flu like symptoms including fever, chills, and fatigue. If left untreated, joint pain, neurological problems, and impaired muscle movement occurs, with further ignorance possibly leading to death. Due to the fact that the symptoms of Lyme Disease (which also include headaches, nausea, and swelling) can also be associated with other diseases such as the flu, Chronic fatigue syndrome, Multiple Sclerosis, and Infectious Arthritis, Lyme disease is often misdiagnosed. In addition, tests used to diagnose Lyme disease are usually unreliable, often giving false negatives during earlier stages of the disease. As a result, thousands are left untreated, causing them to proceed onto the later stages of Lyme disease unknowingly, with the end result being death.
Despite the horrible ending that those with Lyme disease experience, Lyme disease itself is easily curable, with patients only needing antibiotics for 2 weeks until they are healthy again. Therefore, seeing that the only problem with Lyme disease is misdiagnosis, the purpose of this design proposal is to produce a test using synthetically engineered bacteria to increase the accuracy of diagnosing Lyme disease in humans.
Competing Technologies Current tests to diagnose Lyme disease include the ELISA test and the Western blot, however, these tests have a 20%-30% chance of providing a false negative, and sometimes even provide false positives. This is due to the fact that the Borrelia burgdorferi is a very evasive bacteria, as it is a spirochete (has a corkscrew shape). In addition, this bacteria also has a slow reproduction rate, meaning that antibodies may take more time to produce, therefore providing false negatives in the ELISA test and Western blot during earlier stages of Lyme disease.
The Design
Borrelia burgdorferi is a unique bacteria that does not need iron to survive, and has adapted to use only manganese to carry out its daily functions. By creating a bacteria similar to the Leptothrix ochracea that oxidizes iron, the Borrelia burgdorferi can be isolated among other cells. The design begins by modelling how normal gram-negative bacteria receive their iron; an outer membrane receptor detects the Iron in the environment, and periplasmic binding proteins bind to the Iron molecules and transport them to the inner membrane. There, the Iron molecule is then passed on to the Permease proteins, through the inner membrane and to the ATP - binding cassette proteins, where the Iron molecule is then transported to wherever the bacteria needs it. For the second part of the cell, a gene that codes for the oxidization of iron in likeness to the Leptothrix ochracea will be inserted into the bacteria, and will be activated through the promoter in the the presence of iron and either H20, O2, or both. The gene will not terminate unless there is no iron, H20, or 02 in which the inserted bacteria as well as the other surrounding cells will be left to die, leaving the remaining Borrelia burgdorferi in isolation.
The intended results mentioned above when this design is working perfectly is shown in the truth table below.
Iron
H20
02
Iron Oxidization
0
0
0
0
1
0
0
0
1
0
1
1
1
1
0
1
1
1
1
1
0
1
1
0
0
0
1
0
0
1
0
0
For the test itself, cell samples will first be taken from various organs in the body that are suspected to contain Borrelia burgdorferi. Next, the synthetically created bacteria containing the gene to oxidize iron will be added to each of the cell samples. Once inserted with de-oxygenated water, the bacteria will begin to oxidize iron, starving all cells of iron other than the Borrelia burgdorferi, causing them to die off.This process will continue until there is no iron left and all the other cells die, with only the Borrelia burgdorferi bacteria left alive. The dead cells will then be washed away using a Apoptotic Cell Isolation Kit, isolating the Borrelia burgdorferi. Afterwards, a variety of ways can be used to determine if there is isolated bacteria, or if the isolated bacteria is Borrelia burgdorferi, including looking at the sample under a microscope.
Advantages The advantage to this design compared to other technologies is that the test is cheaper than the Western blot test, as the synthetically engineered bacteria will be easy and cheap to produce. In addition, this procedure involves less steps and techniques than both the Western blot and the ELISA test, allowing for more flexibility and allowing for more time to be invested on other patients and cases. Furthermore, this test will be more accurate, as it relies not on the antibodies that the patient produces (as the body is easily tricked into not detecting Borrelia burgdorferi during earlier stages of Lyme disease) but actually isolating and identifying the bacteria itself, eliminating both false positives and false negatives when working perfectly.
Potential Problems It is possible that this test, as well as the synthetically engineered bacteria, will come across several problems in its use. In regards to the bacteria, the amount of bacteria needed is unknown in order to optimize the rate of oxidization of iron, and therefore it may take a very long time for the iron be completely eliminated from that environment. For the test as a whole, the cell samples taken from the patient may not contain the bacteria at all even if it is existent in the organ due to the fact that Borrelia burgdorferi has a slow reproductive rate, rendering the test ineffective. The cell sampling can also contribute to incorrect results; educated guesswork is required to determine which parts of the body the Borrelia burgdorferi could reside in. Furthermore, the Borrelia burgdorferi bacteria will have to be kept alive throughout the duration of this test, possibly by adding manganese and other substances throughout the test. In addition, the synthetically engineered bacteria and other cells that require iron may have other survival mechanisms that may keep these cells alive for longer than expected. Despite these problems, these issue can easily be resolved with further research on Borrelia burgdorferi and the testing of this method to detect Lyme disease.
When I was in Elementary School, I remembered hating Biology and dreading to go to science class. However, after a course in Grade 9 Science with an awe-inspiring teacher who majored in Biology, I learned to love everything from plants to genetics, and was hooked! I'm hoping to major in Biology when I graduate, but I'm still unsure whether I want to attend University in Canada or in the U.S., as both are very worthy contenders!
Iron-Oxidizing Bacteria to Detect Lyme Disease
Introduction/ Purpose
A bacterial virus that affects at least 300,000 Americans a year, Lyme Disease is surprisingly still a large mystery to the public despite the rapidly increasing infection rates. It begins with a bite from a tick that latches onto the human skin. As ticks are small in size, they are often left unnoticed and undisturbed as they are feeding on humans. If the tick is left to feed for over 48 hours, the tick is more likely to transfer Borrelia burgdorferi (the type of bacteria that causes Lyme disease) to its victim; the chances of contracting the disease increases as the tick’s feeding time is prolonged.
An adult female tick. From: https://extension.umaine.edu/ipm/tickid/tick-photos/
The Borrelia burgdorferi bacteria itself is a spirochete, meaning that it has a spiral shaped body and flagella enclosed between the outer and inner membranes. Many other threatening diseases are spirochetes as well, implicating the consequences of contracting Borrelia burgdorferi. Compared to the other members of the spirochete family, the Borrelia burgdorferi is rather small, and its flagella is unique as it is hidden beneath the outer membrane instead of it being external. In addition, it does need iron to go through the electron transport chain to produce energy, and also has a slow reproductive rate, tricking the body into taking a longer time to develop antibodies to defend itself.
Lyme Disease Bacteria (Borrelia burgdorferi) structure. From:https://microbewiki.kenyon.edu/index.php/Borrelia_burgdorferi_NEU2011
Early symptoms of Lyme disease include a bull’s-eye rash, a definitive indicator of Lyme disease, however, many do not have such obvious symptoms and develop more flu like symptoms including fever, chills, and fatigue. If left untreated, joint pain, neurological problems, and impaired muscle movement occurs, with further ignorance possibly leading to death. Due to the fact that the symptoms of Lyme Disease (which also include headaches, nausea, and swelling) can also be associated with other diseases such as the flu, Chronic fatigue syndrome, Multiple Sclerosis, and Infectious Arthritis, Lyme disease is often misdiagnosed. In addition, tests used to diagnose Lyme disease are usually unreliable, often giving false negatives during earlier stages of the disease. As a result, thousands are left untreated, causing them to proceed onto the later stages of Lyme disease unknowingly, with the end result being death.
The bull’s eye rash, a definitive symptom of Lyme disease. From: __http://www.webmd.com/arthritis/ss/slideshow-lyme-disease__
Despite the horrible ending that those with Lyme disease experience, Lyme disease itself is easily curable, with patients only needing antibiotics for 2 weeks until they are healthy again. Therefore, seeing that the only problem with Lyme disease is misdiagnosis, the purpose of this design proposal is to produce a test using synthetically engineered bacteria to increase the accuracy of diagnosing Lyme disease in humans.
Competing Technologies
Current tests to diagnose Lyme disease include the ELISA test and the Western blot, however, these tests have a 20%-30% chance of providing a false negative, and sometimes even provide false positives. This is due to the fact that the Borrelia burgdorferi is a very evasive bacteria, as it is a spirochete (has a corkscrew shape). In addition, this bacteria also has a slow reproduction rate, meaning that antibodies may take more time to produce, therefore providing false negatives in the ELISA test and Western blot during earlier stages of Lyme disease.
The Design
Borrelia burgdorferi is a unique bacteria that does not need iron to survive, and has adapted to use only manganese to carry out its daily functions. By creating a bacteria similar to the Leptothrix ochracea that oxidizes iron, the Borrelia burgdorferi can be isolated among other cells. The design begins by modelling how normal gram-negative bacteria receive their iron; an outer membrane receptor detects the Iron in the environment, and periplasmic binding proteins bind to the Iron molecules and transport them to the inner membrane. There, the Iron molecule is then passed on to the Permease proteins, through the inner membrane and to the ATP - binding cassette proteins, where the Iron molecule is then transported to wherever the bacteria needs it. For the second part of the cell, a gene that codes for the oxidization of iron in likeness to the Leptothrix ochracea will be inserted into the bacteria, and will be activated through the promoter in the the presence of iron and either H20, O2, or both. The gene will not terminate unless there is no iron, H20, or 02 in which the inserted bacteria as well as the other surrounding cells will be left to die, leaving the remaining Borrelia burgdorferi in isolation.
The intended results mentioned above when this design is working perfectly is shown in the truth table below.
For the test itself, cell samples will first be taken from various organs in the body that are suspected to contain Borrelia burgdorferi. Next, the synthetically created bacteria containing the gene to oxidize iron will be added to each of the cell samples. Once inserted with de-oxygenated water, the bacteria will begin to oxidize iron, starving all cells of iron other than the Borrelia burgdorferi, causing them to die off.This process will continue until there is no iron left and all the other cells die, with only the Borrelia burgdorferi bacteria left alive. The dead cells will then be washed away using a Apoptotic Cell Isolation Kit, isolating the Borrelia burgdorferi. Afterwards, a variety of ways can be used to determine if there is isolated bacteria, or if the isolated bacteria is Borrelia burgdorferi, including looking at the sample under a microscope.
Advantages
The advantage to this design compared to other technologies is that the test is cheaper than the Western blot test, as the synthetically engineered bacteria will be easy and cheap to produce. In addition, this procedure involves less steps and techniques than both the Western blot and the ELISA test, allowing for more flexibility and allowing for more time to be invested on other patients and cases. Furthermore, this test will be more accurate, as it relies not on the antibodies that the patient produces (as the body is easily tricked into not detecting Borrelia burgdorferi during earlier stages of Lyme disease) but actually isolating and identifying the bacteria itself, eliminating both false positives and false negatives when working perfectly.
Potential Problems
It is possible that this test, as well as the synthetically engineered bacteria, will come across several problems in its use. In regards to the bacteria, the amount of bacteria needed is unknown in order to optimize the rate of oxidization of iron, and therefore it may take a very long time for the iron be completely eliminated from that environment. For the test as a whole, the cell samples taken from the patient may not contain the bacteria at all even if it is existent in the organ due to the fact that Borrelia burgdorferi has a slow reproductive rate, rendering the test ineffective. The cell sampling can also contribute to incorrect results; educated guesswork is required to determine which parts of the body the Borrelia burgdorferi could reside in. Furthermore, the Borrelia burgdorferi bacteria will have to be kept alive throughout the duration of this test, possibly by adding manganese and other substances throughout the test. In addition, the synthetically engineered bacteria and other cells that require iron may have other survival mechanisms that may keep these cells alive for longer than expected. Despite these problems, these issue can easily be resolved with further research on Borrelia burgdorferi and the testing of this method to detect Lyme disease.